Comparative efficacy of renal preservation solutions to limit functional impairment after warm ischemic injury  N. Ahmad, J.R. Pratt, D.J. Potts, J.P.A. Lodge  Kidney International  Volume 69, Issue 5, Pages 884-893 (March 2006) DOI: 10.1038/sj.ki.5000063 Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 1 Assessment of postischemic urine production. (a) There was a significant (P<0.05) drop in urine osmolality in the postischemic period in all groups from the mean normal level of 300 mOsm kg−1 body weight. While the saline and EC groups exhibited isosthenuria and near isosthenuria respectively, the kidneys in the HOC, PBS140, and UW groups showed progressive concentration of urine. The second-hour urine osmolality in the PBS140 and UW groups was significantly higher than that in both the saline and the EC groups (P<0.05). The urine output (b) in the HOC, PBS140 and UW groups exhibited initial postischemic polyuria. At 2 h postischemia, the urine flow rate in these groups was significantly higher than their respective preischemic control and the second hour urine volume in saline control (P<0.05). The urine flow rate in the EC group was not significantly different from the saline control group in this study, but this indicated severe renal damage. Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 2 Assessment of post ischemic renal function. Inulin clearance may be used as an indicator of GFR. (a) This was significantly reduced at 2 h postischemia compared to preischemic values in all groups (P<0.05), either expressed directly or as a percentage of preischemic values (not shown). GFR measured by inulin clearance is linked to urine flow, of which there was very little in the saline and EC groups. GFR was higher in the HOC, UW, and PBS140 groups, although there were not significant differences within these groups in postischemic inulin clearance. (b) Osmolar excretion rate and (c) sodium excretion rate were also lower in the saline and EC groups compared to the HOC, UW, and PBS140 groups, which exhibited significantly higher osmolar and sodium excretion rates compared to preischemic values (P<0.05). Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 3 Assessment of urinary markers of renal damage. (a) Glucose excretion in the EC group was significantly higher than that detected in any of the other groups which showed low levels of urinary glucose. Glucose is a constituent of EC that probably accounted for the glucose excretion data in the EC flushed group. (b) Data for urinary protein showed low urinary protein in all groups except HOC. (c) This was also reflected in the expression of urinary protein/creatinine ratio. (d, e) Levels of urinary α- and πGST were not detected in the saline and EC groups because of the lack of urine production in this group, (d, e) but data for HOC, UW and PBS140 clearly show highly significant increases (P<0.001) in αGSt and πGST, indicative of tubule damage. Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 4 Assessment of indices of tissue injury. (a) Changes in kidney weight showed that there was a significant increase in all groups of perfused organs except PBS140, indicative of retained fluid or cellular edema in these organs. PBS140-perfused tissues were not significantly different in weight than normal kidney controls. (b) An analysis of inulin clearance related to change in kidney weight showed that low GFR was associated with higher weight, suggesting that renal function was impaired by edema, irrespective of the preservation fluid used. (c–e) Macroscopic examination of the organs in this study showed that groups associated with poor function demonstrated gross medullary congestion, which was less marked in the better-performing groups and near normal in those organs with the best postischemic renal function, that is (c) Severe medullary congestion seen without preischemic flush or with saline flush. (e) Mild to moderate medullary congestion seen with EC or, occasionally, with HOC flush. (d) Normal appearance after PBS140 & UW flush. Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 5 Terminal histology of kidneys flushed with different fluids. Histological analysis was performed on tissues harvested at the termination of each experiment. The arterioles of the saline flushed kidneys (a) exhibited widespread subendothelial infiltration of erythrocytes (EI), while proximal tubules demonstrated feathery degeneration of cytoplasm (FD) and widespread separation of epithelial tight junctions (TJ). In places, tubule degeneration (TD) was evident with cellular material and cell casts present in tubular lumen. (b) Medullary arrays of descending and ascending limbs of Loops of Henle (LoH) as well as collecting ducts are difficult to differentiate owing to parenchymal detachment (PD), while the vessels of the vasa recta were congested with erythrocytes. (c, d) Extravasation of erythrocytes (EE) into renal lumenal spaces was also evident. In the EC group, there was widespread feathery degeneration (FD) of proximal tubules. Arterioles (A) and venules (V) appeared patent, but some degree of tissue shrinkage permitted expansion of extracellular spaces between tubules and vessels. Protein casts were present in distal tubules. In the medulla, there was some degree of parenchymal detachment (PD) in the collecting ducts (CD), and the descending limbs (DL) and ascending limbs (AL) of LoH contained protein casts (PC). Vascular congestion (VC) of the vasa recta was widespread. (e, f) In the HOC group, erythrocyte infiltration (EI) was again a feature in the cortical arterioles. There was also expansion of extracellular spaces as well as a moderate incidence of TJ separation. Elements of the medullary arrays were identifiable with protein casts (PC) in the collecting ducts (CD) vascular congestion in the vessels of the Vasa Recta but with clear descending and ascending limbs of LoH. (a, h) Kidneys of the UW flushed group showed degeneration of tight junctions (TJ) in places, as well as moderate occurrence of edematous tubules (OT). Some signs of tubular degeneration were seen (arrows), but erythrocyte infiltration was not a feature in this group. Medullary arrays appeared clear with only minor indications of congestion. (i, j) The PBS140 flushed group exhibited well-maintained architecture, with only minor occurrence of tubular degeneration or erythrocyte infiltration of vascular smooth muscle cells. Medullary arrays appeared clear of congestion, but showed signs of some dilation. Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions

Figure 6 Comparison of preservation solutions to influence postischemic renal function. The log10 webplot compares the mean data from the HOC, UW, and PBS140 groups against mean preischemic kidney data. Normal control data means are expressed as 1, and experimental mean data were plotted as values relative to normal for all parameters except α- and μGST, which are practically zero in the normal kidney. For these two data sets, the actual experimental values are plotted for the HOC, UW, and PBS140 groups. The closer the experimental values are to the normal value, for each parameter, the better the function of the experimental kidneys and the less severe the tissue injury. This overview of data shows that, for most of the parameters measured, PBS140-perfused kidneys maintained their renal function closer to normal, and the indices of tubular injury were less than in the kidneys in the HOC and UW groups. Taken together, this plot demonstrates that PBS140-preserved kidneys were less damaged than kidneys in the other groups. Kidney International 2006 69, 884-893DOI: (10.1038/sj.ki.5000063) Copyright © 2006 International Society of Nephrology Terms and Conditions